There is a continuous need in medical practice, research and diagnostic procedures for rapid, accurate and qualitative or quantitative determinations of biological substances which are present in biological fluids at low concentrations. For example, the presence of drugs, narcotics, hormones, steroids, polypeptides, prostaglandins or infectious organisms in blood, urine, saliva, vaginal secretions, seminal fluids and other biological fluids has to be determined in an accurate and rapid fashion for suitable diagnosis or treatment.
To provide such determinations, various methods have been devised for isolating and identifying biological substances employing specific binding reactions between the substance to be detected (identified as a "ligand" herein) and compounds specifically reactive with that substance (identified as "receptors" herein). Radioactive or enzyme labels have been used to detect the resulting reactive complex.
One particular type of test which has been developed is what is known in the art as an immunometric or a "sandwich" assay. Such an assay involves "sandwiching" the ligand (such as an antigen) with two or more receptor molecules (such as antibodies) which complex with the compound in a non-interfering manner and at different epitopic sites. Examples of such assays are described in U.S. Pat. No. 4,486,530 (issued Dec. 4, 1984 to David et al) where monoclonal antibodies having high affinity are used. In most sandwich assays, one or more of the receptor molecules are suitably immobilized on an insoluble carrier such as small particles, membranes, plates, or similar objects, as described in U.S. Pat. No. 4,496,654 (issued Jan. 29, 1985 to Katz et al) where a biotinylated antibody is immobilized on an avidin coated support. U.K. Patent No. 2,074,727 describes sandwich assays in which the complex of antigen and antibodies formed is or will be insolubilized at the time of or subsequent to complex formation.
One ligand of particular interest is human chorionic gonadotropin (hCG), a hormone which increases in concentration in a human female soon after conception. It is desirable to measure hCG at the appropriate concentrations in order to detect pregnancy early. A very useful assay and test device for hCG determination is described in copending and commonly assigned U.S. Ser. No. 136,211, filed by Smith-Lewis on Dec. 18, 1987. That assay utilizes a biotinylated antibody to hCG which is immobilized in the test device using an acrylamide polymer. In that assay, as well as others known in the art, a wash solution is used to separate complexed materials from uncomplexed materials.
It has been observed, however, that the contents of the wash solution used can affect background in the test. Unwanted background (that is, detectable species from sources other than hCG) makes it difficult to detect low hCG concentrations. In other words, it reduces assay sensitivity. Moreover, if background is consistently high enough, there may be a high incidence of false results in assays.
It would be highly desirable to have a highly sensitive and rapid test for an immunological ligand, especially hCG, in test specimens which is not prone to high background.